The present invention is directed to a bacteriophage composition useful in treating food products to prevent bacterial contamination by Escherichia coli bacteria.
The Centers for Disease Control and Prevention (CDC) estimates that as many as 33 million people develop food poisoning each yearxe2x80x94about one out of every ten Americansxe2x80x94and about 9,000 die. Since many cases of food poisoning are mistaken for a 24-hour xe2x80x9cstomach flu,xe2x80x9d the actual number of people with foodborn illness is probably much higher. Cerrato, P., xe2x80x9cWhen food is the culprit; food poisoning,xe2x80x9d RN, 62(6):52 (Jun. 1, 1999). The presence of foodborn pathogens in a country""s food supply not only affects the health of the local population, but also represents a potential for spread of pathogens to visitors to the country and to consumers in countries which import food products. Buzby et al., xe2x80x9cEconomic costs and trade impacts of microbial foodborne illness, xe2x80x9d World Health Stat. Q., 50(1-2):57-66 (1997).
Prevention of foodborne illnesses by microbial contamination is of major concern to the food processing industry, regulatory agencies, and consumers. Foodborne microbial contamination occurs both prior to entry into the processing facility, and by cross-contamination in the processing environment. The Food Safety and Inspection Service (FSIS) of the United States Department of Agriculture has instituted new Hazard Analysis and Critical Control Point (HACCP) requirements to reduce the occurrence and number of foodborne pathogens. These regulations must be met by food processors. Although the means of achieving this microbial reduction is left to the discretion of the processor, FSIS expects that antimicrobial treatments will be an important component of HACCP plans. The treatment methods of the present invention, which employ formulations of the bacteriophage compositions of the invention, are useful in meeting the HACCP requirements.
In their efforts to provide a product completely free of microbial contamination, poultry and meat processors have encountered major difficulties in removing microorganisms that infect poultry and meat tissues intended as food products.
E. coli Bacteria
E. coli bacteria are short Gram-negative rods that are part of the normal flora of the intestines of most warm-blooded animals. These organisms are the most common facultative anaerobe in the large bowel and provide protection against colonization by other harmful microbes. There are, though, five distinct groups of E. coli that can cause enteric disease: (1) enteroinvasive, (2) enteropathogenic, (3) enterotoxigenic, (4) enteroadherent, and (5) enterohemorrhagic. Pilot et al., xe2x80x9cThreats from the food we eat; includes related articles; New and Emerging Pathogens, part 3,xe2x80x9d Medical Laboratory Observer, 28:42 (Apr. 1996).
E. coli 0157:H7 bacteria, first described in 1982, is a subset of the enterohemorrhagic variety that produces Shiga-like toxins (verotoxins) 1 and 2. Pilot et al. (Apr. 1996); and xe2x80x9cMedical Experts Urge Radiation of Beef to Kill Deadly Bacteria,xe2x80x9d The New York Times, Jul. 14, 1994, at page A15.
The toxin produced by E. coli 0157:H7 in the intestines can cause anything from a mild diarrhea to severe hemorrhagic colitis, where the cells of the intestinal lining are damaged, allowing blood to pass into stool. In as many as 16% of all cases, the infection progresses to a more serious conditionxe2x80x94hemolytic-uremic syndrome (HUS). Mackenzie, D. L., xe2x80x9cWhen E. coli turns deadly,xe2x80x9d RN, 62(7):28 (Jul. 1, 1999). HUS, which occurs when the bacterial toxin enters the bloodstream through the damaged intestinal wall and travels to the smaller arteries that supply the kidneys, going on to damage those vessels, is characterized by hemolytic anemia, thrombocytopenia, and acute renal failure. In addition, anywhere from 10%-50% of patients with HUS develop long-term sequelae, including cardiomyopathy, pulmonary problems, pericardial effusions, end-stage renal disease, chronic hypertension, hyperglycemia, and encephalopathy. Mackenzie, D. L. (Jul. 1, 1999).
E. coli 0157:H7 is a leading cause of HUS in the United States and a leading cause of acute renal failure in children. Although the overall incidence of E. coli 0157:H7 infection is unknown, estimates based on a 1994 outbreak in the Seattle area suggest that more than 20,000 cases occur in this country each year, and that 250 of them result in death. Mackenzie, D. L. (Jul. 1, 1999); and xe2x80x9cBan the 0157:H7 Bomb,xe2x80x9d Nutrition Action Healthletter, 22:3 (Janurary 1995/Feburary 1995). Young children, the elderly, and people with weakened immune systems are most susceptible to infection and resulting complications.
There is no known treatment for E. coli 0157:H7 infection. Nutrition Action Healthletter, 22:3 (January 1995/Feburary 1995). Antibiotics do little to deter the infection. xe2x80x9cMedical Experts Urge Radiation of Beef to Kill Deadly Bacteria,xe2x80x9d The New York Times, Jul. 14, 1994, at page A15; and Cerrato, P. (Jun. 1, 1999).
One reason this strain of E. coli is so dangerous is that it can cause disease in such low dosesxe2x80x94ingestion of less than 1,000 organisms compared to more than the 10,000,000 needed before the bacterium that causes cholera causes disease. Cerrato, P. (Jun. 1, 1999). Some reports state that E. coli 0157:H7 can cause illness with as few as 69. The New York Times, Jul. 14, 1994, at page A15. As little as one-half cup of E. coli-contaminated cider can cause abdominal pain, bloody diarrhea, vomiting and fever. xe2x80x9cRoadside cider may be risky; may be contaminated with E. coli,xe2x80x9d Environmental Nutrition, 16:8 (September 1993). Such small amounts of disease-causing bacteria cannot be detected by routine meat inspection practices, and the presence of contaminated food products cannot be identified by sight, taste or smell. The New York Times, Jul. 14, 1994, at page A15. Similarly, the pathogen""s acid resistance gives it an advantage over other pathogens. Gastric acid, one of the gastrointestinal tract""s first lines of defense against foodborne illness, has little effect on E. coli 0157:H7. Cerrato, P. (Jun. 1, 1999).
E. coli 0157 is found regularly in the feces of healthy cattle, and is transmitted to humans through contaminated food, water, and direct contact with infected people or animals. Mead et al., xe2x80x9cEscherichia coli 0157:H7,xe2x80x9d The Lancet, 352(9135):1207-1212 (Oct. 10, 1998). Transmission of the infection is primarily linked to consumption of undercooked ground beef, contaminated drinking water, and unpasteurized milk. Hamburger is a major vehicle of foodborne outbreaks of E. coli 0157:H7 infection. Koutkia et al., xe2x80x9cEnterohemorrhagic Escherichia coli 0157:H7,xe2x80x9d American Family Physician, 56:853 (Sep. 1, 1997).
During the slaughter process, intestinal fluid or feces of infected cattle can drip onto the surface of the meat, contaminating it. It is theorized the harmful bacteria on the surface of the raw meat become mixed throughout the meat during the grinding process, where it can better survive the heat of cooking. One hamburger patty can contain the meat from many cows.
Currently, E. coli 0157:H7 is the most common of verotoxin-producing E. coli serotypes. Various sources indicate it comprises from about 60% to more than 90% of all toxin-producing isolates and has been responsible for most outbreaks when food sources have been implicated. Pilot et al. (April. 1996). Distribution of E. coli 0157:H7 probably is worldwide; the majority of cases have been noted in North America and Europe. Pilot et al. (April 1996).
Current Methods of Treating Food Products to Eliminate Bacterial Contamination
A. Irradiation
Food irradiation is the treatment of foods by subjecting them to ionizing radiation, also called ionizing energy. The radiation used in the process comes either from radioactive isotopes of cobalt or cesium or from devices that produce controlled amounts of high-energy electrons, gamma rays, or X rays. The process does not and cannot make the food radioactive. Greenberg et al., xe2x80x9cIrradiated Foods,xe2x80x9d American Counsil on Sci. and Health Booklets, 1-28 (Apr. 30, 1996).
The radiation used to treat foods is called xe2x80x9cionizing radiationxe2x80x9d because it produces ionsxe2x80x94electrically charged particles. Ionizing radiationxe2x80x94including X rays, gamma rays, and beams of high-energy electrons produced by electron acceleratorsxe2x80x94has a higher energy than other, nonionizing radiation such as visible light, television waves, radio waves and microwaves. Greenberg et al. (Apr. 30, 1996).
Food irradiation has been proposed as one solution to the food safety problem. Food irradiation was first approved by the FDA in 1963 for the control of insects in wheat flour and again in 1964 to prevent sprouting in potatoes. Later, it was approved for use in spices, produce, and poultry, and on Dec. 3, 1998, it was approved for beef, lamb, and pork to control disease-causing microorganisms. Klausner, A., xe2x80x9cFood Irradiation: We May Be Zapping Up The Wrong Tree,xe2x80x9d Environmental Nutrition, 17(12):1 (Dec. 31, 1994).
Two radiation sources are practical for food treatment. The first is a tightly sealed metal container of radioactive elementsxe2x80x94cobalt 60 or cesium 137xe2x80x94that produce gamma rays. The rays are directed onto the food being irradiated, with the food itself never being touched by the cobalt or cesium. The second type of radiation source is a machine that produces X rays and high-energy electrons. Neither of these sources has enough energy to make the irradiated foods radioactive. Greenberg et al. (Apr. 30, 1996).
During the irradiation process, food is carried on a conveyor belt into a sealed chamber where it is hit with large doses of energy from a radiation source (usually gamma rays from radio-active cobalt). The energy disrupts molecules in the food, killing insects, molds, fungi and bacteria that cause spoilage, as well as pathogens that cause foodborne illnesses. Klausner, A. (Dec. 31, 1994).
Consumer advocates warn that food irradiation is not a panacea. In fact, they say, it poses safety risks of its own. For example, food irradiation opponents cite research showing the process depletes nutrients (vitamins A, C, and the B vitamin, thiamin). The greater the radiation dose, the greater the losses. In addition, an irradiated food must still undergo normal nutrient-depleting processing, compounding the losses. Klausner, A. (Dec. 31, 1994).
Food irradiation also forms residual substances such as benzene and formaldehydexe2x80x94both known carcinogens. And it produces free radicals and chemically altered food components called radiolytic products, whose long-term health effects are unknown. Klausner, A. (Dec. 31, 1994). It is hypothesized that changes from such food irradiation could lead to more cancers. Id. Moreover, reports on consumer acceptance are mixed. Taste tests for irradiated beef first reported in The Economist failed miserably, as consumers described a xe2x80x9cburnt-hairxe2x80x9d taste. Klausner, A. (Dec. 31, 1994).
Another drawback of irradiation as an bacterial-control process is that irradiation, like heat treatment, does not leave any active agent in an unpackaged food to protect it against reinfestation. Greenberg et al. (Apr. 30, 1996).
B. Chemical Treatments of Meat
Several chemical and physical methods have been proposed to reduce microorganisms in meat products, such as the use of chlorine or chlorine dioxide, ozone, hydrogen peroxide, lactic acid, sodium carbonate, trisodium phosphate, and electrical stimulation. Generally, these methods have shown limited effectiveness in reducing microbial contamination and may affect the physical appearance of the meat products. In addition, such studies show that frequently unacceptably high levels of chemicals are required to kill pathogens such as E. coli 0157:H7. Id.
For example, U.S. Pat. No. 5,366,983, incorporated herein by reference, discloses a method for removing or preventing Salmonella contamination of meat products by treatment with an effective amount of an aqueous solution of a quaternary ammonium cationic surfactant, such as alkylpyridinium, particularly cetylpyridinium chloride (CPC) and cetylpyridinium bromide (CPB).
Disinfestation by irradiation can substitute for some of the former uses of the now-banned fumigant ethylene dibromide (EDB). The possibility of such a replacement is one of the major reasons for renewed interest in irradiation in the United States, as the chemical fumigants being used currently in place of EDB have serious disadvantages, including increased hazards for the workers who must apply them. Greenberg et al. (Apr. 30, 1996).
Another disadvantage of chemical disinfectants is that residues of such chemicals invariably remain on the foodstuffs. Greenberg et al. (Apr. 30, 1996).
Treatment with chemical disinfectants can also produce resistant bacteria. xe2x80x9cGrowing Menace: Antibiotic-Resistant xe2x80x98Supergermsxe2x80x99,xe2x80x9d The Int""l. Council for Health Freedom Newsletter, II(3-4):18 (Fall, 1998); Freeman, C., xe2x80x9cAntimicrobial Resistance: Implications for the Clinician,xe2x80x9d Critical Care Nursing Q., 20(3):21 (Nov. 1997).
C. Pulsed Light
Yet another method for treating food products to eliminate bacterial contamination is treatment with high intensity light emissions, produced by a pulsed power energization technique (PPET). MacGregor et al., xe2x80x9cLight inactivation of food-related pathogenic bacteria using a pulsed power source,xe2x80x9d Letters in Applied Microbiology, 27(2):67-70 (1998). This process comprises dissipating many megawatts (MW) of peak electrical power in the light source in an extremely short energization time (about 1 (mu)s). The light source is subjected to electric field levels greater than could be achieved under conventional continuous operation, which leads to a greater production of the shorter bacteriocidal wavelengths of light.
The usefulness of this process is limited, as the pulsed light emissions can only significantly reduce bacterial populations on exposed surfaces. Such a process is not useful for treating, for example, ground hamburger. This is significant as recent studies indicate that E. coli bacteria can be present on the interior of foodstuffs such as apples, which would not be treatable using a pulsed light process. Buchanan et al., xe2x80x9cContamination of intact apples after immersion in an aqueous environment containing Escherichia coli 0157:H7,xe2x80x9d J Food Prot., 62(5):444-50 (May, 1999).
There is a need in the art for a superior composition and method for treating food stuffs to eliminate bacterial contamination by E. coli bacteria, particularly toxin-producing E. coli bacteria. The present invention satisfies these needs.
The present invention is directed to novel phage compositions useful in treating food products to minimize or eliminate bacterial contamination by E. coli bacteria, particularly toxin-producing E. coli bacteria. The phage compositions can be formulated with suitable carriers.
Also encompassed by the invention are methods of treating food products, it such as meat (including but not limited to poultry, beef, lamb, and pork), juices, spices, and produce.
Both the foregoing general description and the following detailed description are exemplary and explanatory and are intended to provide further explanation of the invention as claimed. Other objects, advantages, and novel features will be readily apparent to those skilled in the art from the following detailed description of the invention.